Disruption of epithelial cell-cell adhesion by exogenous expression of a mutated nonfunctional N-cadherin.

Abstract

Cadherins, a family of transmembrane cell-cell adhesion receptors, require interactions with the cytoskeleton for normal function. To assess the mechanisms of these interactions, we studied the effect of exogenous expression of a mutant N-cadherin, cN390A, on epithelial cell-cell adhesion. The intracellular domain of cN390A was intact but its extracellular domain was largely deleted so that this molecule was not functional for cell adhesion. cDNA of cN390A was attached to the metallothionein promoter, and introduced into the keratinocyte line PAM212 expressing endogenous E-and P-cadherin. When the expression of cN390A was induced by Zn2+, cadherin-dependent adhesion of the transfected cells was inhibited, resulting in the dispersion of cell colonies, although their contacts were maintained under high cell density conditions. In these cultures, cN390A was expressed not only on the free surfaces of the cells but also at cell-cell junctions. The endogenous cadherins were concentrated at cell-cell junctions under normal conditions. As a result of cN390A expression, however, the endogenous cadherins localizing at the cell-cell junctions were largely diminished, suggesting that these molecules were replaced by the mutant molecules at these sites. As a control, we transfected the same cell line with cDNA of a truncated form of Ncadherin whose intracellular C terminus had been deleted leaving the extracellular domain intact. This molecule had no effect on cell-cell adhesion, nor did it localize to cell-cell contact sites. We also found that the association of the endogenous cadherins with a-and 3-catenins and plakoglobin was not affected by the expression of cN390A, which also formed a complex with these molecules, suggesting that no competition occurred between the endogenous and exogenous cadherins for these cytoplasmic proteins. These and other additional results suggest that the nonfunctional cadherins whose intracellular domain is intact occupy the sites where the endogenous cadherins should localize, through interactions with the cytoskeleton, and inhibit the cadherin adhesion system.

Purification and spectroscopic characterization of a recombinant amino‐terminal polypeptide fragment of mouse epithelial cadherin

“…Furthermore, a number of previous observations [14] indicate that other portions of the cadherin molecule are necessary for cell adhesive function. For example, tight cell-cell junctions mediated by cadherin interactions may require anchoring to a complex of cytoskeletal proteins through the cytoplasmic domain and also possibly molecular clustering via lateral interactions in the plasma membrane [33,34]. The present study supports this view in that a single CAD repeat itself does not form a stable dimer or an oligomer.…”

Cdc42 GTPase and Rac1 GTPase act downstream of p120 catenin and require GTP exchange during gastrulation of zebrafish mesoderm

“…p120 catenin and b-catenin are the prototypic members of a small family of armadillo repeat domain proteins (Daniel and Reynolds, 1995;Hatzfeld and Nachtsheim, 1996;Hatzfeld et al, 2003;Gu et al, 2009;Zhao et al, 2011). All p120 catenins and ARVCF bind the intracellular juxtamembrane region of the cytoplasmic tail of cadherins to maintain strong cell-cell adhesion (Fujimori and Takeichi, 1993;Reynolds et al, 1996;Ireton et al, 2002;Davis et al, 2003;Kausalya et al, 2004;Xiao et al, 2007;Ishiyama et al, 2010). There are four members of the p120 catenin gene family in zebrafish: ARVCF, p120 catenin d1, p120 catenin d2a (also known as NPRAP/Neurojungin), and p120 catenin d2b (also known as Delta-catenin, plakophilin 4 (PKP4), and p0071 catenin).…”

Involvement of p120 Carboxy-Terminal Domain in Cadherin Trafficking

“…An investigation of p120-deficient SW48 carcinoma cells showed that restoring p120 expression efficiently rescued proper epithelial morphology by stabilizing E-cadherin and increasing its abundance (Ireton et al, 2002). Previous reports showed that the expression of extracellular domain deleted dominant-negative (DN) cadherins typically downregulated endogenous cadherins (Fujimori and Takeichi, 1993;Kintner, 1992;Nieman et al, 1999;Troxell et al, 1999;Zhu and Watt, 1996). Overexpression of mutated DN-VE-cadherin constructs lacking either the p120 or β-catenin binding sites showed that downregulation of endogenous VE-cadherin was caused by sequestration of p120, but not β-catenin, indicating that p120 plays an essential role in the regulation of cadherin stability (Xiao et al, 2003).…”